Saibil, H.R., Fenton, W.A., Clare, D.K., Horwich, A.L. (2013) Structure and Allostery of the Chaperonin GroEL. J Mol Biol. 2013 May 13;425(9):1476-87
A recent paper, by Professor Helen
Saibil’s team at Birkbeck, reviews the current understanding of chaperonin
GroEL. Chaperonins attract unfortunate
proteins which are incompletely or incorrectly folded and provide them with an
isolated chamber in which to bind and release until they achieve their native
functional state. GroEL and its partner
GroES are profiled in PPS in Section 7 (symmetry)
and Section 8 (action as a chaperone).
GroEL is remarkable in its
construction. It consists of 14
identical protomers arranged
in two back-to back rings, each of the two rings with seven subunits. This forms a barrel with a 7-fold rotational
symmetry axis through its centre and, perpendicular to this, seven 2-fold axes
of symmetry, giving an overall symmetry of 72.
Each subunit comprises two main domains
linked by an intermediate domain (see figure (c)). The largest domain is equatorial at the
centre of the barrel. This contains the
ATP binding site and is in contact with its two neighbours in the ring as well as
the equatorial domains of its partner ring.
These domains form a stable platform from which the other two domains
undergo large movements orchestrated by the cycle of ATP binding, hydrolysis
The apical domains are exposed at the outer
ends of the GroEL barrel. They are
smaller and include a hydrophobic surface which is the binding site for many
different nonnative polypeptides. The
intermediate domain has a hinge at the junction with each of the two main
domains, such that it can mediate large movements of the domains as rigid
bodies. This can be seen by comparing
figures (c) and (f).
X-ray crystal structures of GroEL and
GroEL-GroES complexes. (a) Longitudinal
cross-section of GroEL (PDB 1OEL).
(b) Top view of the GroEL barrel. (c) A protomer of GroEL, aligned
approximately as the top left protomer in (a).
(d-f) Show the same set of views with GroES (d-e) and ATP (f) bound (PDB 1SVT). Example helices have been coloured to
demonstrate the extent of the rotation angles.
The red and orange helices of the apical domains can be seen to undergo
a significant rotation. Compare this
with the relatively minor movements of the green helices in the intermediate
domains and the violet helices of the equatorial domains.
X-ray crystal structures of GroEL and GroEL-GroES complexes. (a) Longitudinal cross-section of GroEL (PDB 1OEL). (b) Top view of the GroEL barrel. (c) A protomer of GroEL, aligned approximately as the top left protomer in (a). (d-f) Show the same set of views with GroES (d-e) and ATP (f) bound (PDB 1SVT). Example helices have been coloured to demonstrate the extent of the rotation angles. The red and orange helices of the apical domains can be seen to undergo a significant rotation. Compare this with the relatively minor movements of the green helices in the intermediate domains and the violet helices of the equatorial domains.
The operations of the GroEL chaperone are initiated by rapid binding of ATP to the equatorial domain of one of the rings. This is followed by the binding of the unstructured, partly folded or misfolded polypeptide.
Natively folded proteins have their hydrophobic residues buried in the stabilising core whilst those which have lost their way have exposed hydrophobic patches. These patches bind to the hydrophobic surfaces of the apical domains.
The final ligand is GroES, a ring of seven homo-oligomers, which forms a lid for the GroEL barrel. Each GroES monomer has a flexible hydrophobic loop which binds to the hydrophobic regions of the apical domain alongside the substrate polypeptide. This loop is visible in figure (d).
The apical domains undergo significant concerted rotations together as one movement, with the domains being held as rigid bodies (compare figures (a) and (d)). These rotations replace the hydrophobic polypeptide binding surface with hydrophilic residues, so propelling the nonnative protein into the central lidded cavity, where it is isolated to refold.
As ATP binding stimulates positive cooperative movements within the cis ring, that is the ring binding the nucleotide, it is also responsible for negative cooperation between the rings. This means that while the movements are coordinated to bind GroES and promote protein folding in the ATP bound cis ring, in the trans or partner ring the opposite rotation prompts the release of GroES, more than 100Å away, and the expulsion of the now native protein.
Mutation studies have revealed that salt bridges, which are studied in PPS Section 9, hold the rings steady until full ATP occupancy is achieved and are probably involved in the positive cooperativity whilst the negative cooperativity is thought to be triggered by a pivoting of the equatorial domains. This interferes with the staggered contacts between each equatorial domain and two of its partner equatorial domains on the opposite ring.
Recent work using single particle cryo-electron microscopy techniques, which is studied in the TSMB course, has captured images of the intermediate states between ATP binding and the active chaperone state where GroES is fully bound.
Once ATP binds, the intermediate and apical domains tilt 35˚ sideways from the lower hinge. This brings the intermediate domain towards the ATP binding pocket where the residue ASP398 forms several hydrogen bonds. This action causes the breakage of salt bridges between the intermediate and apical domains of neighbouring protomers and between neighbouring apical domains with new salt bridges forming which support the new tilted architecture.
Following this, the apical domains lift and separate, to use an old advertising slogan, causing further breakage of salt bridges between apical domains. This separation could help to unfold the misfolded polypeptide before it is released into the chaperone chamber and also positions the hydrophobic binding areas for docking of the GroES binding loops.
Once GroES is bound, the apical domains lift even further outwards and undergo a 100˚ twist to create the active folding chaperone with the GroES lid in a domed position and the polypeptide is released into the cavity to complete its folding.
The next stage is hydrolysis of the ATP, which triggers the release of the ligands on the cis ring, and the acceptance of ligands on the trans ring.
The mechanism is believed to involve separation of a β sheet contact between equatorial domains of the trans ring. The equatorial domains are primarily responsible for holding the rings together so that the ADP complex has reduced stability. Hydrolysis is followed by ATP binding to the equatorial domains of the trans ring. This promotes the pivoting of the equatorial domains that defines negative cooperativity and the discharge of GroES, the native protein and ADP from the cis ring, although the exact movements which lead to the discharge are unknown.
It seems likely that the release is mediated through a reversal of the twist in the cis apical domains. This is speculation, however, as this good Samaritan of nanomachines has not yet given up all of its trade secrets. The progress made to date, however, in large part by Professor Saibil’s team, is a striking demonstration of the power of this recently developed method in structural biology.